A microassay for measuring synaptosomal 3H-dopamine and 3H-metabolite release.
A modified synaptosomal superfusion apparatus is described which uses less than 10 micrograms of tissue per replicate sample and facilitates the routine separation of 3H-DA, 3H-DOPAC, and 3H-HVA. A flow rate of 1.5 ml/min allows superfusion without the use of reuptake or monoamine oxidase inhibitors. Superfusate samples are collected directly onto alumina columns for the separation of 3H-DA and its acid metabolites. Total recovery of authentic 3H-DA applied via superfusion was 87.63(1.10) percent [Mean(SEM)]. Contamination of the acetic acid eluate fraction, containing 80.98(1.15)% of the total DA, by DOPAC and HVA was less than 0.1%. To illustrate the utility of this technique, the relative proportions of 3H-DA and 3H-metabolites released from synaptosomes by 6 mM potassium and 1 microM reserpine were compared.
Petrie
E C
EC
Department of Veterans Affairs Medical Center, Palo Alto, CA 94304.
Lombrozo
L
L
Csernansky
J G
JG
D015102:3,4-Dihydroxyphenylacetic Acid / Q000378:metabolism; D000818:Animals; D002851:Chromatography, High Pressure Liquid; D004298:Dopamine / Q000378:metabolism*; D006719:Homovanillic Acid / Q000378:metabolism; D066298:In Vitro Techniques; D008297:Male; D008832:Microchemistry; D009714:Nucleus Accumbens / Q000378:metabolism; D011188:Potassium / Q000494:pharmacology; D051381:Rats; D011919:Rats, Inbred Strains; D012110:Reserpine / Q000494:pharmacology; D013574:Synaptosomes / Q000378:metabolism*
D015102:3,4-Dihydroxyphenylacetic Acid; D012110:Reserpine; D011188:Potassium; D004298:Dopamine; D006719:Homovanillic Acid